Ail organisms are plagued by pathiogens, wiiicfi can rapidly evolve to overcome iiost defenses, resulting in increased host damage or mortality (virulence). Genetic variability of hosts provides one defense against this deadly potential. Experimental evolution of a murine retrovirus revealed 156-fold increases in fitness and 11- fold increases in virulence after just 10 serial passages through single-genotype hosts, but these dramatic increases were completely abolished when the virus faced five alternating host genotypes. Conditions that promote high transmission are also predicted to favor high virulence. For example, the virulence of Marek's disease virus of chickens has been steadily increasing over the past decades as barriers to transmission and host genetic diversity have been reduced. The long term objective of this study is to identify the mechanisms controlling virulence evolution of pathogens, which will lead to approaches for controlling the diseases emerging from such virulence increases. This study manipulates levels of both transmission and host genetic diversity during experimental viral evolution to quantify how each factor controls the evolution of pathogen transmissibility, replication and virulence (Aim 1). These experiments will be conducted in two hosts - mice (Mus) and chickens (Gallus), representing a model mammal and an avian agricultural species. For each host, two separate viral pathogens will be independently evaluated. We will utilize deep sequencing techniques to interrogate the entire genomes ofthe evolved viruses to identify the genetic basis of viral transmissibility, replication and virulence changes (Aim 2), which will fuel future experiments to discover specific mechanisms. These empirical data will form a basis for the development of mathematical models needed to deduce host consequences from pathogen replication and virulence evolution (Aim 3). The ability to study these processes experimentally and in real time provides a powerfulyet under-utilized tool for dissecting the complex interactions between hosts and pathogens, which will be important for understanding and controlling pathogen-caused diseases. RELEVANCE (See instructions): Beyond infectious disease itself, pathogens cause other diseases such as cancer, autoimmunity, and escape from vaccine immunity. This study identifies the causes of evolving pathogen virulence, which is fundamental to: 1) controlling pathogen-caused diseases, 2) controlling the emergence of new infectious diseases, 3) preparing against the use of pathogens as bioterrorism weapons and 4) controlling the global, antibiotic- resistance crisis bv minimizing the need for orophvlactic antibiotics in domestir animals